SynB3 conjugated QBP1 passes blood-brain barrier models and inhibits polyQ protein aggregation

Background: Polyglutamine diseases are degenerative diseases in the central nervous system caused by CAG trinucleotide repeat expansion which encodes polyglutamine tracts, leading to the misfolding of pathological proteins. Small peptides can be designed to prevent polyglutamine diseases by inhibiting the polyglutamine protein aggregation, for example, polyglutamine binding peptide 1(QBP1). However, the transportation capability of polyglutamine binding peptide 1 across the blood-brain barrier is less efficient. We hypothesized whether its therapeutic effect could be improved by increasing the rate of membrane penetration. Objectives: The objective of the study was to explore whether polyglutamine binding peptide 1 conjugated cell-penetrating peptides could pass through the blood-brain barrier and inhibit the aggregation of polyglutamine proteins. Methods: n order to investigate the toxic effects, we constructed a novel stable inducible PC12 cells to express Huntington protein that either has 11 glutamine repeats or 63 glutamine repeats to mimic wild type and polyglutamine expand Huntington protein, respectively. Both SynB3 and TAT conjugated polyglutamine binding peptide 1 was synthesized, respectively, and we tested their capabilities to pass through a Trans-well system and subsequently studied the counteractive effects on polyglutamine protein aggregation. Results: The conjugation of cell-penetrating peptides to SynB3 and TAT enhanced the transportation of polyglutamine binding peptide 1 across the mono-cell layer and ameliorated polyglutamine-expanded Huntington protein aggregation; moreover, SynB3 showed better delivery efficiency than TAT. Interestingly, it has been observed that polyglutamine binding peptide 1 specifically inhibited polyglutamine-expanded protein aggregation rather than affected other amyloidosis proteins, for example, β-Amyloid. Conclusion: Our study indicated that SynB3 could be an effective carrier for polyglutamine binding peptide 1 distribution through the blood-brain barrier model and ameliorate the formation of polyglutamine inclusions, thus SynB3 conjugated polyglutamine binding peptide 1 could be considered as a therapeutic candidate for polyglutamine diseases.

The MID1 Protein: A Promising Therapeutic Target in Huntington’s Disease

Huntington’s disease (HD) is caused by an expansion mutation of a CAG repeat in exon 1 of the huntingtin (HTT) gene, that encodes an expanded polyglutamine tract in the HTT protein. HD is characterized by progressive psychiatric and cognitive symptoms associated with a progressive movement disorder. HTT is ubiquitously expressed, but the pathological changes caused by the mutation are most prominent in the central nervous system. Since the mutation was discovered, research has mainly focused on the mutant HTT protein. But what if the polyglutamine protein is not the only cause of the neurotoxicity? Recent studies show that the mutant RNA transcript is also involved in cellular dysfunction. Here we discuss the abnormal interaction of the mutant HTT transcript with a protein complex containing the MID1 protein. MID1 aberrantly binds to CAG repeats and this binding increases with CAG repeat length. Since MID1 is a translation regulator, association of the MID1 complex stimulates translation of mutant HTT mRNA, resulting in an overproduction of polyglutamine protein. Thus, blocking the interaction between MID1 and mutant HTT mRNA is a promising therapeutic approach. Additionally, we show that MID1 expression in the brain of both HD patients and HD mice is aberrantly increased. This finding further supports the concept of blocking the interaction between MID1 and mutant HTT mRNA to counteract mutant HTT translation as a valuable therapeutic strategy. In line, recent studies in which either compounds affecting the assembly of the MID1 complex or molecules targeting HTT RNA, show promising results.

A discontinuous Galerkin model for fluorescence loss in photobleaching of intracellular polyglutamine protein aggregates

Background: Intracellular phase separation and aggregation of proteins with extended poly-glutamine (polyQ) stretches are hallmarks of various age-associated neurodegenerative diseases. Progress in our understanding of such processes heavily relies on quantitative fluorescence imaging of suitably tagged proteins. Fluorescence loss in photobleaching (FLIP) is particularly well-suited to study the dynamics of protein aggregation in cellular models of Chorea Huntington and other polyQ diseases, as FLIP gives access to the full spatio-temporal profile of intensity changes in the cell geometry. In contrast to other methods, also dim aggregates become visible during time evolution of fluorescence loss in cellular compartments. However, methods for computational analysis of FLIP data are sparse, and transport models for estimation of transport and diffusion parameters from experimental FLIP sequences are missing. Results: In this paper, we present a computational method for analysis of FLIP imaging experiments of intracellular polyglutamine protein aggregates also called inclusion bodies (IBs). By this method, we are able to determine the diffusion constant and nuclear membrane permeability coefficients of polyQ proteins as well as the exchange rates between aggregates and the cytoplasm. Our method is based on a reaction-diffusion multi-compartment model defined on a mesh obtained by segmentation of the cell images from the FLIP sequence. The discontinuous Galerkin (DG) method is used for numerical implementation of our model in FEniCS, which greatly reduces the computing time. The method is applied to representative experimental FLIP sequences, and consistent estimates of all transport parameters are obtained. Conclusions: By directly estimating the transport parameters from live-cell image sequences using our new computational FLIP approach surprisingly fast exchange dynamics of mutant Huntingtin between cytoplasm and dim IBs could be revealed. This is likely relevant also for other polyQ diseases. Thus, our method allows for quantifying protein dynamics at different stages of the protein aggregation process in cellular models of neurodegeneration.